Glutathione Synthetase Research Articles

In vitro cytotoxicity of resin cement and its influence on the expression of antioxidant genes

Aim: This study evaluated cytotoxicity and antioxidant gene expression of resin cements on human gingival fibroblasts (hGF). Materials and Method: RelyX UltimateTM(RXU), VariolinkTMII(VLII), and RelyXU200TM(RXU200) resin cements were incubated with culture medium for 24 h to obtain eluates. Then, the eluates were applied over hGF to assess cell viability at 24 h, 48 h, and 72 h and antioxidant gene expression at 24 h. hGF cultures non-exposed to the eluates were used as Control. Data were submitted to ANOVA and Bonferroni tests (α≤0.05). Results: RXU and RXU200 reduced the number of viable cells in 24 h. Longer exposure to cement extracts caused cell death. Gene expression showed peroxiredoxin 1 (PRDX1) induction by all resin cement types, and superoxide dismutase 1 (SOD1) induction by RXU200 and VLII. Moreover, RXU200 induced not only PRDX1 and SOD1, but also glutathione peroxidase 1 (GPX1), catalase (CAT), and glutathione synthetase (GSS). Conclusions: All resin cements showed toxicity, and induced antioxidant genes in hGF. Antioxidant gene induction is at least partly associated with cytotoxicity of tested cements to oxidative stress experience.

Propofol suppresses OGD/R-induced ferroptosis in neurons by inhibiting the HIF-1α/YTHDF1/BECN1 axis

ABSTRACT Background Ischemia/reperfusion (I/R) is a pathological process that causes severe damage. Propofol is known to alleviate I/R-related injury; however, the exact function and underlying mechanisms are not fully understood. Methods Using an oxygen glucose deprivation/re-oxygenation (OGD/R) method, an in vitro I/R injury model was induced. The cell viability and the level of Fe2+, glutathione synthetase (GSH), and malondialdehyde (MDA) were evaluated using kits. Luciferase reporter gene assay, chromatin immunoprecipitation, and RNA immunoprecipitation (RIP) were used to verify the interaction between molecules. The m6A level of BECN1 mRNA was determined through methylated RIP. Results Propofol-treated OGD/R models showed reduced levels of Fe2+ and MDA, while the cell viability and the level of GSH increased. Propofol inhibited ferroptosis by down-regulating HIF-1α in OGD/R-treated HT22 cells. HIF-1α is bound to the promoter region of YTHDF1 to promote its transcription, and YTHDF1 promoted ferroptosis by stabilizing the mRNA of BECN1. The suppressive effect of propofol on OGD/R-induced ferroptosis was reversed by the overexpression of YTHDF1. Conclusions Our study revealed that the HIF-1α/YTHDF1/BECN1 axis in OGD/R-treated HT22 cells promotes ferroptosis, and administration of propofol can inhibit this axis to avoid cell death. This study provides a novel insight for the neuroprotective function of propofol.

Extracellular Production of Glutathione by Recombinant Escherichia coli K-12

The goal of this study was to produce a sufficient amount of glutathione in the fermentation medium without the addition of cysteine. This would simplify and reduce the cost of its purification. In addition to reducing the cost of cysteine, it also avoids the inhibition of bacterial growth by cysteine. The gshA, gshB, and cysE genes of Escherichia coli were cloned under the control of the strong T5 promoter of the pQE-80L plasmid and introduced into an E. coli strain knocked out for the genes encoding γ-glutamyltranspeptidase and the GsiABCD glutathione transporter, which are responsible for the recycling of excreted glutathione. The overexpression of the gshA and gshB genes, genes for γ-glutamylcysteine synthetase and glutathione synthetase, and the cysEV95R D96P gene, a gene for serine acetyltransferase with the V95R D96P mutation that makes it insensitive to cysteine, were effective on glutathione production. Na2S2O3 was a good sulfur source for glutathione production, while the addition of Na2SO4 did not affect the glutathione production. With the addition of 50 mM glutamic acid and 75 mM glycine, but without the addition of cysteine, to the simplified SM1 medium, 4.6 mM and 0.56 mM of the reduced and oxidized glutathione, respectively, were accumulated in the extracellular space after 36 h of batch culture. This can eliminate the need to extract glutathione from the bacterial cells for purification.

A Comparative Analysis of Chemical Constituents and Antioxidant Effects of Dendrobium fimbriatum Hook Fractions with Different Polarities.

The aim of this study was to investigate the chemical composition and antioxidant capacity of various polar fractions obtained from Dendrobium fimbriatum Hook (DH). First, a 90% ethanol-aqueous extract of DH (CF) was subjected to sequential fractionation using different organic solvents, resulting in the isolation of a methylene chloride fraction (DF), an ethyl acetate fraction (EF), an n-butanol fraction (BF), and a remaining water fraction (WF) after condensation. Additionally, the CF was also subjected to column chromatography via a D101 macroreticular resin column, eluted with ethanol-aqueous solution to yield six fractions (0%, 20%, 40%, 60%, 80%, and 100%). UPLC-Q-Exactive Orbitrap-MS/MS analysis identified a total of 47 chemical compounds from these polar fractions, including fatty acids, amino acids, phenolic acids, flavonoids, organic heterocyclic molecules, and aromatic compounds. Moreover, DF, EF, and the 60%, 80%, and 100% ethanol-aqueous fractions had higher total phenol content (TPC) and total flavonoid content (TFC) values and greater 2,2'-azinobis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS-) and 1,1-diphenyl-2-picrylhydrazyl (DPPH)-scavenging abilities. In H2O2-induced HepG2 cells, the aforementioned fractions could increase the activities of antioxidative enzymes NAD(P)H: quinone oxidoreductase 1 (NQO1), superoxide dismutase (SOD), heme oxygenase-1 (HO-1) and catalase (CAT), stimulate glutathione (GSH) synthesis by increasing the activities of glutamic acid cysteine ligase (GCL) and glutathione synthetase (GS), regulate GSH metabolism by increasing glutathione peroxidase (GSH-Px) and glutathione reductase (GR) activities, and reduce levels of reactive oxygen species (ROS) and malondialdehyde (MDA). Furthermore, the antioxidative stress effect of the DH fractions was found to be positively correlated with the activation of nuclear factor-erythroid 2-related factor 2 (Nrf2) protein and the presence of antioxidative chemical constituents. In conclusion, this study highlights the efficacy of both liquid-liquid extraction and macroporous resin purification techniques in the enrichment of bioactive compounds from natural food resources. The comprehensive analysis of chemical constituents and antioxidant effects of different polar fractions from Dendrobium fimbriatum Hook contributes to the understanding of its potential application in functional foods and nutraceuticals.

Transcriptomics and cell painting analysis reveals molecular and morphological features associated with fed-batch production performance in CHO recombinant clones.

Stable, highly productive mammalian cells are critical for manufacturing affordable and effective biological medicines. Establishing a rational design of optimal biotherapeutic expression systems requires understanding how cells support the high demand for efficient biologics production. To that end, we performed transcriptomics and high-throughput imaging studies to identify putative genes and morphological features that underpin differences in antibody productivity among clones from a Chinese hamster ovary cell line. During log phase growth, we found that the expression of genes involved in biological processes related to cellular morphology varied significantly between clones with high specific productivity (qP > 35 pg/cell/day) and low specific productivity (qP < 20 pg/cell/day). At Day 10 of a fed-batch production run, near peak viable cell density, differences in gene expression related to metabolism, epigenetic regulation, and proliferation became prominent. Furthermore, we identified a subset of genes whose expression predicted overall productivity, including glutathione synthetase (Gss) and lactate dehydrogenase A (LDHA). Finally, we demonstrated the feasibility of cell painting coupled with high-throughput imaging to assess the morphological properties of intracellular organelles in relation to growth and productivity in fed-batch production. Our efforts lay the groundwork for systematic elucidation of clone performance using a multiomics approach that can guide future process design strategies.

Structure-based drug designing against Leishmania donovani using docking and molecular dynamics simulation studies: exploring glutathione synthetase as a drug target

In the pursuit of developing novel anti-leishmanial agents, we conducted an extensive computational study to screen inhibitors from the FDA-approved ZINC database against Leishmania donovani glutathione synthetase. The three-dimensional structure of Leishmania donovani glutathione synthetase was constructed by homology modeling, using the crystallographic structure of Trypanosoma brucei glutathione synthetase as a template. Subsequently, molecular docking studies were carried out for a large number of compounds using AutoDock Vina. Among the screened compounds, we selected the top five with strong binding affinity to Leishmania donovani glutathione synthetase but having a very low affinity to its human homolog. Further investigations on protein-ligand complexes were done by conducting molecular dynamics (MD) simulation and MM/PBSA analysis. The results revealed that Olysio (Simeprevir) exhibited the lowest binding energy (-89.21 kcal/mol), followed by Telithromycin (-45.34 kcal/mol). These findings showed that these compounds have the potential to act as inhibitors of glutathione synthetase. Hence, our study provides valuable insights for the development of a novel therapeutic strategy against Leishmania donovani by targeting the glutathione synthetase enzyme. Communicated by Ramaswamy H. Sarma

Single Nucleotide Polymorphism in Glutathione Synthetase Gene in Indian Population with Type 2 Diabetes

Introduction: Our earlier study has confirmed the presence of significantly low glutathione (GSH) in individuals with type 2 diabetes. Low levels of GSH in addition to other factors can be due to the presence of single nucleotide polymorphism (SNP) in the gene glutathione synthetase (GSS). We have therefore checked the presence of SNPs in GSS, known to be associated with GSH deficiency, in the Indian population. Methods: Blood samples were collected from cases (people with diabetes) and controls (individuals without diabetes) for GSH estimation and DNA isolation. Amplicons obtained against selected exons of GSS after polymerase chain reaction amplification were sequenced and analyzed for the presence of SNPs. Results: A significantly low GSH was found in the case compared to control group (P &lt; 0.001). However, we did not find any reported or novel SNPs associated with GSH deficiency in either of the groups. Conclusion: Data obtained suggest that the lower GSH found in individuals with diabetes of Indian origin is not contributed by SNPs in GSS reported otherwise in different population.

Elemene induces cell apoptosis via inhibiting glutathione synthesis in lung adenocarcinoma.

The rhizome of Curcuma wenyujin Y.H. Chen & C. Ling, also known as Wen-E-Zhu, has been used for cancer treatment since ancient times, with roots dating back to the Song Dynasty. Elemene (EE), a sesquiterpene extract with potent anticancer properties, is extracted from Wen-E-Zhu, with β-elemene (BE) being its main active compound, along with trace amounts of β-caryophyllene (BC), γ-elemene and δ-elemene isomers. EE has demonstrated broad-spectrum anti-cancer effects and is commonly used in clinical treatments for various types of malignant cancers, including lung cancer. Studies have shown that EE can arrest the cell cycle, inhibit cancer cell proliferation, and induce apoptosis and autophagy. However, the exact mechanism of its anti-lung cancer activity remains unclear and requires further research and investigation. In this study, the possible mechanism of EE and its main active components, BE and BC, against lung adenocarcinoma was investigated by using A549 and PC9 cell lines. The subcutaneous tumor model of nude mice was constructed to evaluate the efficacy of EE in vivo, then the in vitro half-inhibitory concentration (IC50) of EE and its main active components, BE and BC, on A549 and PC9 cells at different concentrations were determined by CCK-8. Flow cytometry was used to detect the apoptosis and cycle of A549 and PC9 cells treated with different concentrations of BE and BC for 24h. Non-targeted metabolomics analysis was performed on A549cells to explore potential target pathways, which were subsequently verified through kit detection and western blot analysis. Injection of EE in A549 tumor-bearing mice effectively suppressed cancer growth in vivo. The IC50 of EE and its main active components, BE and BC, was around 60μg/mL. Flow cytometry analysis showed that BE and BC blocked the G2/M and S phases of lung adenocarcinoma cells and induced apoptosis, leading to a significant reduction in mitochondrial membrane potential (MMP). Results from non-targeted metabolomics analysis indicated that the glutathione metabolism pathway in A549cells was altered after treatment with the active components. Kit detection revealed a decrease in glutathione (GSH) levels and an increase in the levels of oxidized glutathione (GSSG) and reactive oxygen (ROS). Supplementation of GSH reduced the inhibitory activity of the active components on lung cancer and also decreased the ROS content of cells. Analysis of glutathione synthesis-related proteins showed a decrease in the expression of glutaminase, cystine/glutamate reverse transporter (SLC7A11), and glutathione synthase (GS), while the expression of glutamate cysteine ligase modified subunit (GCLM) was increased. In the apoptosis-related pathway, Bax protein and cleaved caspase-9/caspase-9 ratio were up-regulated and Bcl-2 protein was down-regulated. EE, BE, and BC showed significant inhibitory effects on the growth of lung adenocarcinoma cells, and the mechanism of action was linked to the glutathione system. By down-regulating the expression of proteins related to GSH synthesis, EE and its main active components BE and BC disrupted the cellular redox system and thereby promoted cell apoptosis.

Kidney bean protein hydrolysate as a fish meal replacer: Effects on growth, digestive enzymes, metabolic functions, immune-antioxidant parameters and their related gene expression, intestinal and muscular gene expression

A feeding trial for 70 days was designated on Nile tilapia, Oreochromis niloticus (O. niloticus). A total of 200 fish (34.38 ± 0.05 g) were assigned into five groups, four replicates for each (10 fish/replicate; 40 fish/group) and received five isonitrogenous and isocaloric diets with five replacement levels of fishmeal (FM) with kidney bean (Phaseolus vulgaris L) protein hydrolysate (KBH) (0, 25, 50, 75, 100%). The findings demonstrated a significant increase in the performance metrics (final body weight, total weight gain, average daily weight gain, protein efficiency ratio, and specific growth rate) with a considerable improvement in the feed conversion ratio across all replacement levels, with the greatest value obtained in the KBH75 and KBH100 groups. Regarding the broken line regression analysis, the optimum FM replacement level with KBH was 76%. By replacing FM with KBH, the growth hormone and digestive enzymes (amylase and protease) levels were significantly increased, whilst the glucose and leptin hormone levels significantly declined. The amino acid transporters (solute carrier family 15 member 2 and solute carrier family 26 member 6), insulin growth factor-1, the immune-associated genes (interleukin-8, toll-like receptors-7, and cc-chemokine), and the antioxidant-associated genes (glutathione peroxidase, superoxide dismutase, and Glutathione synthetase) expression was substantially boosted in all replacement levels, where the KBH50 group recorded the highest expression. As well as the mRNA expression of myostatin was down-turned in all replacement levels relative to the KBH0 group. Improving the immune functions (lysozyme, complement 3, nitric oxide, phagocytic %, and phagocytic index) was noticed by FM replacement with KBH. The cumulative mortality as a result of challenging with Aeromonas sobria at the end of the feeding trial was decreased by increasing the replacement level (80, 40, 37, 30, and 25% for KBH0, KBH25, KBH50, KBH75, and KBH100, respectively). From the economic point of view, the feed costs and feed cost/ kg gain declined in the KBH75 and KBH100 diets. From the aforementioned outcomes, KBH could replace the FM up to 100% with improving the growth and their related genes expression, digestive enzymes activity, and immune/antioxidant parameters and their encoding gene expression, and the optimal replacement level was 76%. The replacement of FM with KBH was economically efficient.

Myocardial Glutathione Synthase and TRXIP Expression Are Significantly Elevated in Hypertension and Diabetes: Influence of Stress on Antioxidant Pathways.

Antioxidant protection is one of the key reactions of cardiomyocytes (CMCs) in response to myocardial damage of various origins. The thioredoxin interacting protein (TXNIP) is an inhibitor of thioredoxin (TXN). Over the recent few years, TXNIP has received significant attention due to its wide range of functions in energy metabolism. In the present work, we studied the features of the redox-thiol systems, in particular, the amount of TXNIP and glutathione synthetase (GS) as markers of oxidative damage to CMCs and antioxidant protection, respectively. This study was carried out on 38-week-old Wistar-Kyoto rats with insulin-dependent diabetes mellitus (DM) induced by streptozotocin, on 38- and 57-week-old hypertensive SHR rats and on a model of combined hypertension and DM (38-week-old SHR rats with DM). It was found that the amount of TXNIP increased in 57-week-old SHR rats, in diabetic rats and in SHR rats with DM. In 38-week-old SHR rats, the expression of TXNIP significantly decreased. The expression of GS was significantly higher compared with the controls in 57-week-old SHR rats, in DM rats and in the case of the combination of hypertension and DM. The obtained data show that myocardial damage caused by DM and hypertension are accompanied by the activation of oxidative stress and antioxidant protection.

Alliin enriched standardized fraction of Allium hookeri regulates the glucose homeostasis via upregulating hepatic glutathione pool

IntroductionAllium hookeri is a source of organosulfur compounds including alliin which is an antihyperglycemic phytochemical. PurposeThis study tested the effect of methanolic extract (ME) of A. hookeri on glucose uptake and glutathione biosynthesis using sodium palmitate (0.75 mM) induced insulin resistant cultured hepatocytes. MethodThe plant extract was prepared following standard protocol followed by bioactivity- guided fractionation. The plant extract and fractions were tested for glucose uptake assay, cellular glutathione level. Further, immunoblotting study was carried out to check the protein expression of the genes involved in glutathione biosynthesis and glucose metabolism. ResultsBioactivity-guided fractionation showed that the aqueous fraction from the methanolic A. hookeri extract (ME) was rich in alliin and the alliin-enriched standardized fraction (EFr) appeared to regulate glucose homeostasis as the cellular glutathione pool in the palmitate treated hepatocytes remained unaltered. Alteration in cellular glutathione level affects glucose homeostasis. We also observed that expressions of glutamate cysteine ligase (GCLC), glutathione reductase (GR), genes involved in hepatic glutathione biosynthesis were downregulated in palmitate and upregulated in EFr treated hepatocytes. GLUT2 protein expression was upregulated in the palmitate treated and downregulated in the EFr treated hepatocytes, but no change was observed in the protein expression of the gene glutathione synthase (GS). ConclusionsOur results show that EFr of A. hookeri leaves may be a potent botanical drug against type2 diabetes due to its role in the regulation of hepatic glutathione pool.

Engineered symbiotic bacteria interfering Nosema redox system inhibit microsporidia parasitism in honeybees

Nosema ceranae is an intracellular parasite invading the midgut of honeybees, which causes serious nosemosis implicated in honeybee colony losses worldwide. The core gut microbiota is involved in protecting against parasitism, and the genetically engineering of the native gut symbionts provides a novel and efficient way to fight pathogens. Here, using laboratory-generated bees mono-associated with gut members, we find that Snodgrassella alvi inhibit microsporidia proliferation, potentially via the stimulation of host oxidant-mediated immune response. Accordingly, N. ceranae employs the thioredoxin and glutathione systems to defend against oxidative stress and maintain a balanced redox equilibrium, which is essential for the infection process. We knock down the gene expression using nanoparticle-mediated RNA interference, which targets the γ-glutamyl-cysteine synthetase and thioredoxin reductase genes of microsporidia. It significantly reduces the spore load, confirming the importance of the antioxidant mechanism for the intracellular invasion of the N. ceranae parasite. Finally, we genetically modify the symbiotic S. alvi to deliver dsRNA corresponding to the genes involved in the redox system of the microsporidia. The engineered S. alvi induces RNA interference and represses parasite gene expression, thereby inhibits the parasitism significantly. Specifically, N. ceranae is most suppressed by the recombinant strain corresponding to the glutathione synthetase or by a mixture of bacteria expressing variable dsRNA. Our findings extend our previous understanding of the protection of gut symbionts against N. ceranae and provide a symbiont-mediated RNAi system for inhibiting microsporidia infection in honeybees.

Cystine/Glutamate Xc- Antiporter Induction Compensates for Transsulfuration Pathway Repression by 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) to Ensure Cysteine for Hepatic Glutathione Biosynthesis.

Exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) has been associated with the induction of oxidative stress and the progression of steatosis to steatohepatitis with fibrosis. It also disrupts metabolic pathways including one-carbon metabolism (OCM) and the transsulfuration pathway with possible consequences on glutathione (GSH) levels. In this study, complementary RNAseq and metabolomics data were integrated to examine the hepatic transsulfuration pathway and glutathione biosynthesis in mice following treatment with TCDD every 4 days for 28 days. TCDD dose-dependently repressed hepatic cystathionine β-synthase (CBS) and cystathionine γ-lyase (CTH) mRNA and protein levels. Reduced CBS and CTH levels are also correlated with dose-dependent decreases in hepatic extract hydrogen sulfide (H2S). In contrast, cysteine levels increased consistent with the induction of Slc7a11, which encodes for the cystine/glutamate Xc- antiporter. Cotreatment of primary hepatocytes with sulfasalazine, a cystine/glutamate Xc- antiporter inhibitor, decreased labeled cysteine incorporation into GSH with a corresponding increase in TCDD cytotoxicity. Although reduced and oxidized GSH levels were unchanged following treatment due to the induction of GSH/GSSG efflux transporter by TCDD, the GSH:GSSG ratio decreased and global protein S-glutathionylation levels in liver extracts increased in response to oxidative stress along with the induction of glutamate-cysteine ligase catalytic subunit (Gclc), glutathione synthetase (Gss), glutathione disulfide reductase (Gsr), and glutathione transferase π (Gstp). Furthermore, levels of ophthalmic acid, a biomarker of oxidative stress indicating GSH consumption, were also increased. Collectively, the data suggest that increased cystine transport due to cystine/glutamate Xc- antiporter induction compensated for decreased cysteine production following repression of the transsulfuration pathway to support GSH synthesis in response to TCDD-induced oxidative stress.

Protein Antioxidant Complex of Water Extract of Larvae of Black Beetles &lt;i&gt;Ulomoides dermestoides&lt;/i&gt;

An aqueous extract from the larvae of black beetles Ulomoides dermestoides grown under controlled conditions was obtained and its proteomic analysis based on one-dimensional electrophoretic separation of proteins, their trypsinolysis in a gel and subsequent chromatography-mass spectrometry was performed. Protein identification was performed using MaxQuant v.1.6.3.4 software. The Uniprot database was used to identify proteins by homology. Catalase activity was determined spectrophotometrically by hydrogen peroxide concentration decrease. Antioxidant activity was evaluated by luminol chemiluminescence quenching. The extract contained proteins that can determine the biological activity of the extract: universal regulators of cellular processes calmodulin (62%), cytochrome c-2 (13.5%), nucleoside diphosphate kinase (11.1%), the enzymatic antioxidants superoxide dismutase, catalase, glutathione S-transferase, peroxiredoxin, glutathione synthetase as well as thioredoxin, 70 and 60 kDa heat shock proteins, chitinase complex (13.4% in total). Catalase activity was 6.3 ± 1.1 µmol H2O2/min/mg protein; antioxidant activity of 1 mg protein per ml extract was equivalent to 1.36 ± 0.3 mM trolox. The prospects for practical use of the extract as a natural antioxidant complex were noted.

Glutathione synthetase from Hydra vulgaris: Molecular cloning, overexpression, purification and partial characterization

Glutathione synthetase from Hydra vulgaris: Molecular cloning, overexpression, purification and partial characterization

Radix Paeoniae Alba attenuates Radix Bupleuri-induced hepatotoxicity by modulating gut microbiota to alleviate the inhibition of saikosaponins on glutathione synthetase

Radix Bupleuri (RB) is commonly used to treat depression, but it can also lead to hepatotoxicity after long-term use. In many anti-depression prescriptions, RB is often used in combination with Radix Paeoniae Alba (RPA) as an herb pair. However, whether RPA can alleviate RB-induced hepatotoxicity remain unclear. In this work, the results confirmed that RB had a dose-dependent antidepressant effect, but the optimal antidepressant dose caused hepatotoxicity. Notably, RPA effectively reversed RB-induced hepatotoxicity. Afterward, the mechanism of RB-induced hepatotoxicity was confirmed. The results showed that saikosaponin A and saikosaponin D could inhibit GSH synthase (GSS) activity in the liver, and further cause liver injury through oxidative stress and nuclear factor kappa B (NF-κB)/NOD-like receptor thermal protein domain associated protein 3 (NLRP3) pathway. Furthermore, the mechanisms by which RPA attenuates RB-induced hepatotoxicity were investigated. The results demonstrated that RPA increased the abundance of intestinal bacteria with glycosidase activity, thereby promoting the conversion of saikosaponins to saikogenins in vivo. Different from saikosaponin A and saikosaponin D, which are directly combined with GSS as an inhibitor, their deglycosylation conversion products saikogenin F and saikogenin G exhibited no GSS binding activity. Based on this, RPA can alleviate the inhibitory effect of saikosaponins on GSS activity to reshape the liver redox balance and further reverse the RB-induced liver inflammatory response by the NF-κB/NLRP3 pathway. In conclusion, the present study suggests that promoting the conversion of saikosaponins by modulating gut microbiota to attenuate the inhibition of GSS is the potential mechanism by which RPA prevents RB-induced hepatotoxicity.

Neem seed protein hydrolysate as a fishmeal substitute in Nile tilapia: Effects on antioxidant/immune pathway, growth, amino acid transporters-related gene expression, and Aeromonas veronii resistance

Whereas fishmeal (FM) is a significant protein supply, its expensive cost has limited its diet use. Therefore, it is necessary to find affordable and accessible FM replacers. The potential of neem seed protein hydrolysate (NSPH) to replace FM protein in the Oreochromis niloticus, Nile tilapia diet was studied after two-month feeding trial. Fish (N = 200) of 27.17 ± 0.09 body weight were haphazardly distributed into five groups and fed on experimental diets having five substitution percentages with NSPH: 0% (control), 10%, 20%, 30%, and 40% (NSPH0, NSPH10, NSPH20, NSPH30, and NSPH40, respectively). Following the feeding experiment, the fish were challenged with Aeromonas veronii (1 × 107 CFU/mL) as a pathogenic bacterium. The results revealed that the growth parameters, growth hormone, digestive enzymes, and immune parameters (nitric oxide, complement 3, lysozyme, and phagocytic activity %) values were linearly (p < 0.01) augmented with all NSPH replacement groups as compared with NSPH0 group. Moreover, the NSPH40 group showed the highest expression of muscular insulin-like growth factor 1 and intestinal amino acids and peptide transporters genes, followed by the NSPH30 group, then the NSPH10 and NSPH20 groups. The splenic immune-related genes (phosphoinositide-3-kinase regulatory subunit 3b, toll-like receptors (TLR-2 and TLR-7), transforming growth factor-beta, interleukins (IL-10 and IL-1β), and tumor necrosis factor-alpha) and antioxidant-related genes (glutathione peroxidase and glutathione synthetase) more up-regulated in the NSPH groups with a concentration-dependent manner than the control one. Among A. veronii-infected fish, mortality rate was decreased with increasing NSPH replacement, where the NSPH40 group had the highest significant (p < 0.05) survival rate (83.33%). From overall results, NSPH can substitute 40% of the dietary FM protein in O. niloticus with promoting the fish's immune competence, growth rate, digestive, and absorptive capability, and disease resistance.

The impact of cadmium stress on the ascorbate-glutathione pathway and ascorbate regeneration in tea plants

Ascorbic acid (AsA) and glutathione (GSH) contribute to defense responses under abiotic stresses. The present study explored the ascorbate-glutathione cycle and ascorbate regeneration under high concentration (30 mM) of cadmium in the tea plant (Camellia sinensis L.). The tea leaves showed speckles and necrosis from the third day of Cd treatment. The content of superoxide anion (O2.-) and hydrogen peroxide (H2O2) in the leaves were significantly higher until the seventh day after Cd treatment. The content of O2.- and H2O2 were the highest on the fifth day (212.7 and 153.6 % of the control, respectively). The AsA content increased (86.9 %) on the first day after Cd treatment and decreased significantly in the subsequent days, while GSH showed a reverse trend. The enzymatic activity assays showed that dehydroascorbate reductase (DHAR) and glutathione reductase (GR) involved in AsA regeneration were downregulated considerably after Cd foliar application. In contrast, the activities of ascorbate peroxidase (APX) and monodehydroascorbate reductase (MDHAR) increased on the first day and then declined. Reverse-transcription quantitative PCR showed upregulation of glutathione synthetase (CsGSHS), γ-glutamylcysteine synthetase (Csγ-ECS), and CsMDHAR of the AsA regeneration pathway and downregulation of CsDHAR and CsGR. The expressions of GDP-L-galactose phosphorylase (CsGGP), L-galactose-1-phosphate phosphatase (CsGPP), and L-galactono- 1,4-lactone dehydrogenase (CsGaILDH) of the L-galactose pathway were also downregulated. The results indicated that AsA, which can respond to Cd stress of plants by increasing antioxidant ability, was consumed to scavenge ROS; moreover, Cd stress inhibited AsA synthesis and regeneration, which made that tea plants suffering severe damage.

In silico Characterization of Toxicophores Found in Lemongrass (Cymbopogon citratus) and its Molecular Interaction with Kidney and Liver Enzymes

BACKGROUND OF THE STUDY: Cymbopogon citratus (lemongrass) is known for its various therapeutic claims. However, the abuse and misuse of herbal medicine is not uncommon. This study investigates the potential toxicophores present in lemongrass plant extract and their binding interaction with kidney and liver enzymes in silico. MATERIALS AND METHODS: Phytochemicals were identified from 74 articles from various databases. The toxic substructures, oral bioavailability, and clearance of these phytochemicals were characterized virtually. The binding affinity of these phytochemicals to different liver and kidney enzymes, namely gamma-glutamyl transferase, glutathione S-transferase, glutathione synthase, renin, human alkaline phosphatase, and 1-alpha-hydroxylase, was predicted and visualized. RESULTS: There were twenty distinct phytochemicals present in lemongrass leaf extract identified from various research articles. Of these twenty phytochemicals, only five were determined to be both hepatotoxic and nephrotoxic despite the majority being found to have high oral bioavailability and low liver and renal clearance. Moreover, isoorientin, which has high liver and renal clearance, was the only phytochemical with a high binding affinity to the liver enzyme gamma-glutamyl transferase and kidney enzyme renin. Meanwhile, the rest of the phytochemicals did not demonstrate high binding affinity to the other liver and kidney enzymes, despite having low clearance. CONCLUSION: Overall, these findings suggest that 13 of the phytochemicals identified may not affect the selected enzymes but may influence other liver and kidney enzymes due to their low liver and renal clearance.

Effect of Methyl Jasmonate on the Gene Expression, Encoding Non-Protein Thiol Enzymes in Wheat under Cadmium.

The effect of the phytohormone methyl jasmonate (MJ) on the expression of the TaGS1 and TaPCS1 genes encoding glutathione synthetase and phytochelatin synthase, respectively, which are key enzymes in the synthesis of glutathione and phytochelatins in wheat (cv. Moskovskaya 39), was studied. It was shown for the first time that pretreatment of plants by exogenous MJ (1 μM) leads to an increase in the accumulation of transcripts of the TaGS1 and TaPCS1 genes in leaves without cadmium effect. When cadmium (CdSO4, 100 μM) was added into the nutrient solution, the transcript level of the TaGS1 gene in the plants pretreated by MJ increased (compared to the untreated plants), whereas the transcript level of TaPCS1 remained unaffected. The pretreatment of plants with MJ leads to a lower accumulation of cadmium in the roots and leaves of wheat. At the same time, MJ had no effect on the linear growth parameters of plants, but had a positive effect on the accumulation of biomass under cadmium. It was assumed that MJ is involved in plant tolerance to cadmium by increasing the expression of the TaGS1 and TaPCS1 genes and, as a result, enhancing the synthesis of chelating compounds, as well as by reducing the supply of metal ions to plants.